The goal of the proposed research is to develop a platform technology for simultaneous detection of multiple analytes with better efficiency and lower cost. Currently, immunoassays are extensively used in biomedical research, clinical diagnostics, food/water safety testing, and biological warfare defense. Although they can be performed in a microplate (e.g. 96 wells) to increase sample throughput, the immunoassay in each well is generally limited to one analyte per test. Such a format is inefficient and not cost-effective for the requirement to detect multiple analytes in a sample. For instance, a spectrum of toxic agents should be simultaneously monitored in foods and drinking water. Similarly, a set of biomarkers, rather than an individual biomarker, should be detected for a particular disease to enhance the accuracy. Therefore, a platform to detect multiple analytes simultaneously is desirable for better efficiency and lower cost. The specific aims of the proposed research are to (1) design and optimize an array of microfluidic valves that are required for containing reagents, directing and regulating flows; (2) demonstrate the microfluidics-enabled multiplexed immunoassay array in the format of sandwich assays and study the effects of surface modification, flow rate, reagent concentration on the assay; and (3) validate the immunoassay array by detecting multiple analytes including toxins and biomarkers. The significance of the research lies in the following aspects. First, the proposed research will lead to a platform for simultaneous detection of multiple analytes. Such a capability will lead to efficient and cost- effective methods. Secondly, the microfluidic-enabled immunoassay array will have shorter analysis time and a reduction in the consumption of required sample and reagents. These advantages could have a significant impact since immunoassays are widely used in chemical, biological and clinical laboratories. Thirdly, the proposed research will result in a manufacturable process that allows mass production of low-cost, disposable devices. The ability to be disposable after a single use could have a tremendous impact to applications where cross-contamination of sequential samples is of concern.